Peptide-bound major histocompatibility complex class I molecules associate with tapasin before dissociation from...

Area of Science:

• Immunology
• Molecular Biology
• Cell Biology

Background:

• Major histocompatibility complex (MHC) class I molecules present peptides to CD8 T cells.
• Peptide transport into the endoplasmic reticulum is mediated by the transporter associated with antigen processing (TAP).
• Tapasin (TAP-A) is involved in MHC class I assembly and peptide loading, but its precise function and murine analogue were unknown.

Purpose of the Study:

• To clone and characterize the mouse analogue of tapasin.
• To investigate the role of tapasin in the assembly and peptide loading of MHC class I molecules.
• To compare mouse tapasin with its human counterpart.

Main Methods:

• Cloning of mouse tapasin.
• Amino acid sequence analysis and comparison with human tapasin.
• Coprecipitation assays using anti-TAP1/2, anti-tapasin, and anti-calreticulin antibodies.
• Experiments with TAP2-mutated RMA-S cells and crosslinker-modified peptides.

Main Results:

• The mouse tapasin analogue was cloned, showing high sequence identity to human tapasin, particularly in conserved functional domains.
• Mouse tapasin binds to both TAP1/2 and MHC class I, similar to human tapasin.
• Tapasin associates with TAP1 and peptide-bound MHC class I, suggesting a role in later stages of MHC class I assembly than calreticulin.
• Tapasin appears to control peptide loading onto MHC class I molecules.

Conclusions:

• A functional murine tapasin analogue has been identified.
• Mouse tapasin plays a critical role in the peptide loading of MHC class I molecules within the endoplasmic reticulum.
• Tapasin functions downstream of calreticulin in the MHC class I assembly pathway.